Abstract:

The present invention relates to multi-substituted m-diarylanilines or
pharmaceutically acceptable salts thereof, wherein X, R1-R7 are
as defined in the claims, their preparation process, pharmaceutical
compositions comprising them and their use for the manufacture of a
medicament for anti-HIV.
##STR00001##

8. A process for preparing a compound as claimed in claim 1: ##STR00014##
wherein R1-R7 and X are as defined in the formula
(I),comprising: reacting substituted m-dihalobenzene of formula (II) with
p-substituted anilines in the presence of a strong base to form N-aryl
substituted aniline of formula (IV); coupling the intermediate of formula
(IV) and multi-substituted phenols or anilines, or heating in the
presence of Pd catalyst or coupling under microwave condition to produce
m-di(aromatic)-polysubstituted anilines of formula (I).

9. A pharmaceutical composition comprising a compound as claimed in claim
1 or a pharmaceutically acceptable salt thereof and one or more
pharmaceutically acceptable carriers or excipients.

10. Use of a compound as claimed in claim 1 or a pharmaceutically
acceptable salt thereof for the manufacture of a medicament for treating
HIV infection associated diseases or disorders.

Description:

TECHNICAL FIELD

[0001]The invention relates to a class of multi-substituted
m-diarylaniline derivatives with anti-HIV activity, its preparation
process, a pharmaceutical composition comprising the same, and their use
for the manufacture of a medicament for anti-HIV.

BACKGROUND ART

[0002]Human immunodeficiency virus (HIV) is a RNA virus. The surface of
this virus is a layer of double lipid membrane. The membrane encapsulates
2 single chain RNA and some important enzymes (e.g. reverse
transcriptase, protease, integrase) and structural protein (p24, p17, p7,
etc). The membrane surface of the virus has two very important
glycoproteins gp120 and gp41. The gp120 is outside of the membrane, gp41
bestrides the double lipid membrane and forms a composite with gp120.
Their main function is to recognize and attack cells having CD4 surface
receptors in human immune system, such as lymphocyte (T cell),
macrophage. HIV cannot be propagated in vitro, it can be replicated and
regenerated only by means of human cells. The replication course of HIV
can be approximately divided into following stages: binding and fusing of
virus and host cells, reverse transcription of virus genes, integration,
transcription and translation, and assembly and release of virus. HIV
continuously replicates in such a circulation course, infects human
immune cells, destroys human immune system, and finally causes the
complete loss of human immune function, so that patients are placed in
the risk of various infections without resistance ability. Theoretically,
only if a drug can interrupt any link during the replication of virus,
the purpose of inhibiting virus and treating diseases can be achieved.

[0003]Up to now, there are 30 kinds of chemical drugs and their
combinations that have been approved to be marketed for clinic anti-HIV
infection and treating AIDS. Existing drugs are divided into five classes
in terms of action mechanism: nucleoside reverse transcriptase inhibitors
(NRTIs), non-nucleoside reverse transcriptase inhibitors (NNRTIs),
protease inhibitors (PIs), fusion inhibitors (Enfuvirtide), and entrance
inhibitors (Maraviroc). The use of existing drugs alone or in combination
can effectively inhibit the replication of virus, however, the main
problem commonly faced is drug resistance. HIV viruses may exhibit some
mutation after interacting with drugs. Mutated virus cannot be inhibited
by drugs, and still continuously replicates a great deal of virus in vivo
as before drug administration. Therefore, seeking and developing a new
generation of anti-HIV drugs having new structure type, new action
mechanism, new action target or exhibiting strong inhibition effect on
virus having drug resistance has been recently a focus in drug study
field.

[0004]At present, there are only three public listed non-nucleoside
reverse transcriptase inhibitors (Nevirapine, Delavirdine, Efavirenz).
These drugs have advantages of structural diversity, high efficiency and
low toxicity, clear target and action mechanism as well as
non-competitive inhibitors, and occupy important places in anti HIV
combination therapy (HAART). However, their main problem is to cause drug
resistance easily. In order to overcome the disadvantages of the existing
drugs, it is needed to seek a new generation of non-nucleoside reverse
transcriptase inhibitors drugs which can effectively inhibit the
replication of wild type and multiple drug resistance HIV virus strains.

SUMMARY OF THE INVENTION

[0005]The inventor has found in the study of new anti-HIV drugs that a
class of compounds having novel structure have rather strong anti-HIV
activity and may be developed to be new anti-HIV drugs.

[0006]Therefore, the present invention relates to, at first aspect, a
compound of multi-substituted m-diarylanilines represented by formula (I)
or a pharmaceutically acceptable salt thereof:

[0016]The present invention relates to, at second aspect, a process for
preparing a compound of formula (I) or a pharmaceutically acceptable salt
thereof.

[0017]The present invention relates to, at third aspect, a pharmaceutical
composition comprising at least one compound of formula (I) or a
pharmaceutically acceptable salt thereof and one or more pharmaceutically
acceptable carriers or excipients.

[0018]The present invention relates to, at fourth aspect, a use of a
compound of formula (I) or a pharmaceutically acceptable salt thereof for
the manufacture of a medicament for treating HIV infection associated
diseases or disorders.

[0019]The term "alkyl radical" used in the present invention includes
alkyl, alkenyl and alkynyl.

[0020]The term "5-membered heteroaryl" used in the present invention means
5-membered heteroaromatic ring system containing 1-3 heteroatoms selected
O, S or N, including but not being limited to furyl, pyrrolyl, thienyl,
pyrazolyl, etc.

[0021]The term "5-membered heteroaryl having on the ring structure thereof
aldehyde, ketone, cyano, α,β-unsaturated cyano, alkenyl,
alkynyl, aldehyde or ketone group substitution" mentioned in the present
invention can be shown as follows, including but not being limited to:

[0099]The compounds of the present invention can be prepared through
following reaction routes:

##STR00006##

[0100]wherein R1-R7 and X are as defined in the formula (I).

[0101]Substituted m-dihalobenzene of formula (II) is allowed to react with
p-substituted anilines in the presence of a strong base to form N-aryl
substituted aniline, i.e. intermediate of formula (IV); then the
intermediate of formula (IV) and multi-substituted phenols or anilines
are subjected to coupling reaction, or heated in the presence of Pd
catalyst or coupled under microwave condition to produce the target
compound of multi-substituted m-diarylanilines as shown in formula (I).

[0102]Besides, a part of the target compounds of formula (I) are obtained
by two-step coupling reaction simultaneously under microwave reaction
condition, i.e. "one pot reaction".

[0103]For example, the compound of formula (Ia) according to the present
invention can be prepared by following synthetic routes:

##STR00007##

[0104]wherein, R1, R2, R3 and R7 are as defined in the
formula (I); the compound of formula (II) having appropriate R1,
R2, R3 substituents can be selected as starting materials based
on requirements.

[0105]Arylamine Coupling Synthesis of Intermediate of Formula (IV)

[0106]Particularly, m-dihalo-nitrobenzene (II) and p-substituted aniline
or substituted phenol (formula IIIa) are allowed to react at a
temperature of between room temperature and 130° C. for from 5
minutes to 24 hours in the presence of potassium tert-butoxide, sodium
hydride, triethylamine, pyridine, N,N-dimethylaminopyridine, or potassium
carbonate/cuprous halide (I) in a solvent of DMF, acetonitrile, THF or
DMSO to give N-(p-substituted aryl) aniline or phenylate intermediate
(IVa). The molar ratio of reactant II/III is 1:1.1-1:2.

[0107]This coupling reaction can also be carried out at a temperature of
150-180° C. under microwave condition in a solvent of DMF or DMSO
for 10-30 minutes to give an intermediate (IV-a). The molar ratio of base
to reactant is as defined above.

[0108]Synthesis of Target Compound (I)

[0109]For example, when X in the formula is --O--, its synthetic method is
as follows:

[0110]Method 1: N-aryl-m-chloroaniline (formula IV-a) and
multi-substituted phenol (formula V) are heated in an atmosphere of
nitrogen to 130° C. and allowed to react in the presence of Cu or
cuprous reagent (e.g. CuI, CuBr) catalyst and K2CO3 in a
solvent of DMSO, DMF for about 4 hours.

[0111]Method 2: In dried ethyl ether, tri-substituted phenol of formula V
is allowed to react with NaH to form sodium salt, and then the sodium
salt and N-aryl-m-chloroaniline of formula (IV-a) are refluxed in DMF for
0.5-8 hours.

[0112]Method 3: The intermediate of formula (IV-a) is allowed to react
with sodium phenolate or potassium phenolate under microwave condition
for 5-10 minutes.

[0113]Method 4: The intermediate of formula (IV-a) is allowed to react
with substituted phenol (formula V) and potassium carbonate under
microwave condition in a solvent of DMSO for 5-10 minutes.

[0114]Also, for example, when the X in the target compound formula is
--NH--, its synthetic method is as follows:

##STR00008##

[0115]wherein each substituent is as defined in the formula (I); the
compound of formula (IV) having appropriate R1, R2, R3
substituents can be selected as starting materials based on requirements.

[0116]Method 5: The intermediate chlorobenzene of formula (IV) is allowed
to react with substituted aniline of formula (VI) at 140-160° C.
in an atmosphere of nitrogen in the presence of Cu, CuI or CuBr catalyst
and K2CO3 in an aprotic polar solvent (e.g. DMSO or DMF) for
about 6 hours.

[0117]Method 6: The intermediate chlorobenzene of formula (IV) is allowed
to react with substituted aniline of formula (VI) at about 100° C.
in an atmosphere of nitrogen in the presence of Pd catalyst and
Cs2CO3 in toluene solvent for 1-20 hours.

[0118]Method 7: If the arylamine of formula (VI) is liquid, no other
solvent is required, the arylamine of formula (VI) can be directly mixed
with the intermediate of formula (IV) and allowed to react at the boiling
point temperature of arylamine under microwave for about 15-20 minutes.

[0119]Method 8: The arylamine of formula (VI) is allowed to react with the
intermediate chlorobenzene of formula (IV) at a molar ratio of 4:1 at a
temperature of 200-250° C. under microwave in the presence of
potassium tert-butoxide in DMSO or NMP solvent for 15-20 minutes.

[0120]Similarly, the compound of formula (I) wherein X has other
definitions can be prepared too.

[0121]The multi-substituted m-diarylanilines shown in formula (I)
according to the present invention exhibit strong anti-HIV activity and
high selectivity in the cell experiments of inhibiting HIV replication
(MT-2 and H9 lymphocyte). Hence, further study on the compounds of the
present invention is helpful to develop new anti-HIV drugs.

[0123]According to the present invention, the compounds of formula (I) of
the present invention can faun pharmaceutical compositions with
pharmaceutically acceptable customary carriers and excipients. The
pharmaceutical compositions can be administered orally or
non-intestinally. The pharmaceutical composition of the present invention
can be prepared by conventional methods in this art into various dosage
forms, including but being not limited to: tablets, capsules, solutions
or suspensions, granulates or injections, which are administered orally
non-intestinally.

[0124]Further, it should be pointed out that the application dosage and
application method of the compounds of the present invention depend on
various factors, including age, body weight, sex, natural health
condition and nutrition state of patients, the activity of the compounds
applied, administration time, metabolism rate, order of severity of
conditions and doctor's subjective judgment. The application dosage is
preferably between 0.01 and 100 mg/kg body weight/day.

EMBODIMENTS

[0125]Following examples are used to further explain the present
invention. However, the present invention is not limited by the examples
at any way.

[0151]Reference document (J. Med. Chem. 2004, 47, 756-760). Lymphocyte H9
was cultured at 37° C. in culture medium 1640 and 5% CO2. The
tested compound was initially dissolved in DMSO, and then diluted with
culture medium to conventional screening concentration: 100, 20, 4, 0.8
μg/mL. The cultured H9 cells were divided into two parts, wherein one
of the parts was infected with HIV virus (IIIB) (m.i.o. 0.1-0.01
infectious Units/cell), which was used for assaying activity. The other
part of the cells was not added with viruses but only with culture
medium, which was used for assaying toxicity. The two parts of the cells
were cultured under completely identical conditions (37° C., 5%
CO2) for 4 hours, washed with fresh culture medium for three times,
and then were added respectively to the prepared tested samples of
different concentrations or to culture medium (the latter was positive
infection control or negative drug control), and meanwhile AZT was used
as positive drug control. All these cells were cultured at 37° C.
in 5% CO2 for 4 days. In the fourth day, the cell membranes of the
cells infected by viruses were removed, and the cytosol was tested by P24
antigen ELISA method to assay the activity of the sample, expressed using
EC50. EC50 was an effective concentration when virus
replication was inhibited by 50%. The part of the cells without viruses
added were tested by the method of counting cells to assay the toxicity
of the sample, expressed using CC50. CC50 was a concentration
when growth cells were killed by 50%.

Example 22

Assay of Anti-HIV Activity (MT-2 Cell Model)

[0152]Reference documents (Jiang, S., et al. Antimicrob. Agents Chemother.
2004, 48, 4349-4359). In 96 wells culture plate, 50 μl of different
concentrations of compound solutions were mixed with HIV-1IIIB virus
strains (100 TCID50) in the same volume, incubated at 37° C.
for 30 minutes, added with 100 μl TM-2 cell (1×105/mL, RPIM
1640 culture medium containing 10% serum), mixed uniformly, and incubated
at 37° C. overnight. In the second day, 150 μl supernatant was
sucked off and supplemented with fresh culture medium in the same volume,
the compound solutions were incubated continuously at 37° C. for 3
days, and the cell pathology effect (CPE) was recorded in the fourth day.
Then 100 μl cultured supernatant was sucked up to determine p24
antigen therein with 5% Triton X-100 lytic virus particles by using ELISA
method. Briefly, HIVIG (2 μg/ml) was used to encapsulate enzyme
standard plate, sealed with 1% fat-free milk, and then added with virus
lytic solution, incubated at 37° C. for 60 minutes. After the
plate was washed completely, anti-p24 monoclonal antibody-183-12H-5C,
goat anti-mouse antibody labeled with biotin and horse radish peroxidase
labeled with avidin were added. Then the compounds were developed with
TMB, and the light density was tested at 450 nm. The virus inhibition
concentration of 50% (EC50) of the compounds was calculated by using
CalcuSyn software.

Example 23

Cytotoxicity Test of Compound

[0153]Reference documents (Jiang, S., et al. Antimicrob. Agents Chemother.
2004, 48, 4349-4359). In 96 wells culture plate, 50 μl of different
concentrations of compound solutions were mixed with PBS in the same
volume, incubated at 37° C. for 30 minutes, added with 100 μL
TM-2 (or H9) cell (1×105/mL, RPIM 1640 culture medium
containing 10% serum), mixed uniformly, and incubated at 37° C.
overnight. In the second day, 150 μL supernatant was sucked off and
supplemented with fresh culture medium in the same volume, the compound
solutions were incubated continuously at 37° C. for 3 days, and
added in the fourth day with 50 μL fresh XTT solution (1 mg/mL)
containing PMS, after 4 hours later, the light density of the compounds
was determined at 450 nm The cytotoxicity concentration of 50%
(CC50) of the compounds was calculated by using CalcuSyn software.

[0155]The results have shown that the compounds of formula I according to
the present invention are anti-HIV active compounds having new skeletal
structure. Since this class of compounds exhibit good molecular
flexibility, the compounds of the present invention may have strong
inhibition activity to HIV tolerant virus strains in large extent and may
be developed as new anti-HIV drugs having novel structure.